The reliability and efficacy of communication between neurons is governed by the fundamental properties of synapses. In addition, modification of synaptic properties is likely to be a crucial component of learning and formation of memories. The long-term objectives of this research program are to understand the elementary steps in release of neurotransmitter from synaptic terminals, and how they can be modulated. Although many of the molecules in the presynaptic terminal have been identified, their precise roles in various steps in release and recycling of vesicles are incompletely understood. Synapses contain functionally heterogeneous populations of vesicles. Release is thought to occur from a small pool of vesicles termed the readily releasable pool, and when this pool is fully or partially depleted, it is filled with vesicles from the reserve pool. This work will focus on characterizing the different pool of synaptic vesicles, the movement of vesicles between them and activity-dependent changes in the characteristics of the vesicle pools. Experiments are designed to use the fluorescent dyes and optical microscopy, combined with molecular biology and electrophysiology, to study synaptic vesicle release and trafficking at visualized individual synapses. The specific objectives are to determine (i) the mechanisms regulating the size and accessibility of the recycling vesicle pool, (ii) the roles of peripherally associated synaptic vesicle proteins in trafficking of vesicles between the different pools, and (iii) the mechanisms in activity-dependent, long-term changes in the size of the vesicle pools. Since synaptic transmission is adversely affected in many disorders of the nervous system, a better understanding of synaptic release mechanisms will provide a rational basis for treatment.